Insulin action at the cellular level is initiated by binding of the hormone to specific glycoprotein receptor sites on the surface membrane of the cell. These receptors have been shown to consist of two major subunits with Mr = 135K (Alpha) and Mr = 95K (Beta). The Alpha subunit appears to contain the insulin binding site. Recently, we and others have demonstrated that the Beta subunit undergoes a phosphorylation reaction, and have presented evidence that this subunit is a tyrosine-specific protein kinase. The present proposal requests support to characterize this property of the receptor and investigate its role in the mechanism of insulin action. This may be of great importance, since in diabetes and many other disorders of carbohydrate metabolism, there appears to be an alteration in insulin action at the cellular level. Specific experiments will be conducted to characterize the protein kinase activity of the receptor in in vitro and in vivo systems. This will include kinetics of reaction, phosphoamino acid determinations, and peptide mapping of the phosphorylation sites. The insulin receptor itself is a tyrosine kinase, but in intact cells is phosphorylated at both tyrosine and serine. Thus, we will attempt to identify and purify the serin kinase involved in this reaction. The effect of the receptor kinase will be investigated using cellular enzymes and proteins involved in insulin action, and attempts will be made to identify the natural substrates of the receptor. Studies will also be conducted to determine the role of receptor phosphorylation in receptor function and to study cell mutants altered in this regard. The receptor kinase will be characterized by radiation inactivation and the phosphorylation sites sequenced. These studies should provide a basis for understanding this reaction. In addition, these studies may help elucidate the mechanism of insulin action and provide important new information on the pathogenesis of diabetes.